Epstein-Barr virus (EBV) is a human herpesvirus that infects both B lymphocytes and epithelial cells. Depending upon cell type and differentiation state, infection leads to lytic replication with cell death or latent infection. Latent infections are associated with several epithelial- and B-cell malignancies. By identifying key cellular factors and pathways regulating EBV latency and lytic replication and drugs that affect their activities, we hope to develop lytic induction therapies for treating patients with EBV-associated cancers. The immediate- early BZLF1 and BRLF1 genes of EBV encode multifunctional proteins essential for lytic replication. While transcription from their promoters, Zp and Rp, respectively, is strongly silenced in latently infected cells, cell differentiation and factors activated by signaling pathways can induce their expression, leading to reactivation into lytic replication. We have identifled several physiologically relevant factors that play key roles in regulating this switch: ZEB1, ZEB2, p53, Oct-1, Oct-2, Smads via TGF-p signaling, and the Zp ZIIR element. We have preliminary data indicating important roles as well for HIF-1 a, Pax5, and Blimp-la and have identified 2 drugs, nufiin-3 and L-mimosine, that induce EBV reactivation via effects on some of these factors. We have developed a mouse model with a humanized immune system that can be use to examine factors affecting EBV-induced tumorigenesis and effects of drugs on growth and killing of EBV+ lymphomas. We have also identified a non-tumorigenic epithelial cell line, NOKs, in which we can study effects of differentiation state on EBV's life cycle. Here, we propose to determine: (i) how HIF-1 a, the ZEBs, PAX5, and Blimp-la play key roles in controlling EBV's latent-lytic balance in cells in culture, during differentiation of EBV+ NOKs and B cells, and in humanized mice; and (ii) whether nufiin-3 or L-mimosine together with HDAC inhibitors can efficiently reactivate EBV, leading to highly selective killing of some types of EBV+ cells lines in culture and tumors in humanized mice, especially when used in combination with ganciclovir, a nucleoside analog prodrug activated by a virus-encoded kinase. The findings obtained from these experiments will identify key cellular factors and pathways that regulate the balance between latency and lytic replication of EBV in epithelial and B cells, may identify effective lytic-inducing drugs for treating EBV-associated cancers, and may lead to a candidate vaccine for immunization against EBV by identifying an EBV mutant unable to establish viral latency. Drs. Kenney and Mertz collaborate extensively, including holding bimonthly joint research group meetings and co-authoring publications in which both labs made key contributions.